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United States Patent | 5,324,667 |
Macri | June 28, 1994 |
The present invention relates to a method for detecting fetal Down syndrome (Trisomy 21), trisomy 13, trisomy 18 and other chromosomal anomalies during prenatal screening by analyzing blood samples from a pregnant woman. More particularly the present invention relates to a method for improving detection efficiency in screening for the anomalies by measuring the amount of the free beta human chorionic gonadotropin (HCG) and nicked or fragmented or aberrant forms of free beta (HCG), all of which are referenced throughout this application as free beta (HCG) in blood samples from pregnant women.
Inventors: | Macri; James N. (170 Sidney St., Oyster Bay, NY 11771) |
Appl. No.: | 925844 |
Filed: | August 7, 1992 |
Current U.S. Class: | 436/518; 435/7.9; 435/7.92; 436/65; 436/86; 436/87; 436/510; 436/548 |
Intern'l Class: | G01N 033/49; G01N 033/493 |
Field of Search: | 436/501,510,518,817,818,87,86 435/7.92,7.9 |
4016250 | Apr., 1977 | Saxena | 424/1. |
4123224 | Oct., 1978 | Givner et al. | 422/59. |
4565687 | Jan., 1986 | Khazaeli et al. | 424/1. |
4874693 | Oct., 1989 | Bogart | 435/7. |
5037305 | Aug., 1991 | Aleck | 434/262. |
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54-126723 | Oct., 1979 | JP | . |
WO89/00696 | Jan., 1989 | WO | . |
WO90/08325 | Jul., 1990 | WO | . |
Arab et al.--Biological Abstracts report of meeting held Oct. 12-15, 1988, New Orleans, La. (39th Annual Meeting of Am. Soc. of Human Genetics). International Search Report from European Patent Office, mailed May 15, 1990, in PCT/US 90/00291 (based on U.S. Appl. Ser. No. 07/420,775 and others). Am. J. Human Genetics (Suppl. ), vol. 43, No. 3, Sep. 1988, H. Arab et al.: "Maternal serum beta human chorionic gonadotropin . . . ". Prenatal Diagnosis, vol. 7, 1987, Mark H. Bogart et al. "Abnormal maternal serum chorionic . . . ". British Med. Journal, vol. 297, Oct. 1988, N. J. Wald et al.: "Maternal serum screening . . . ". Patent Abstracts of Japan, vol. 3, No. 143, C 66, abstract of JP 54-126723, publ. 1979-02-10, Teikoku Zoki Seiy Aku K.K. American Journal of Obstetrics and Gynecology, vol. 48, No. 7, 1984, I. R. Merkatz et al.: "An association between low maternal serum . . . ". Ann. Endocrinol. (Paris) vol. 45(4-5), pp. 269-280, 1984, Gaspard, U. et al., "HCG and its Subunits in Normal and Pathological Pregnancies". ACTA Endocrinol.; 100(1), pp. 109-113, 1982, L'Hermite-Baleriaux, M. et al., "Alteration of free hCG Subunit Secretions in Ectopic Pregnancy". Am. J. Human Genetics, vol. 43, No. 3, Sep. 1988, Bharathur et al.: "Amniotic fluid beta hCG levels Associated with Down syndrome and Other Chromosome Abnormalities". Conference Poster, Society of Perinatal Obsetricians, Feb. 7-8, 1992, Rotmensch et al., "Peptide Heterogeneity of Human Chorionic Gonadotropin (hCG) and Its .beta.-Subunit in Down Syndrome Pregnancies". The Lancet, Oct. 8, 1988, p. 851, Cuckle et al., "First-Trimester Biochemical Screening For Down Syndrome". Prenatal Diagnosis, vol. 10, 245-251 (1990), Brock et al., "First-Trimester Maternal Serum Biochemical Indicators in Down Syndrome". The Yale Journal of Biology and Medicine; 64(1991), 627-637, Cole et al., "The Biological and Clinical Significance of Nicks in Human Chorionic Gonadotropin and Its Free .beta.-Subunit". Clin. Chem. 38/1, 26-33 (1992), Kardana et al., "Polypeptide Nicks Cause Erroneous Results in Assays of Human Chorionic Gonadotropin Free .beta.-Subunit". Clin. Chem. 37/6, 809-814 (1991), Spencer, K. "Evaluation of an Assay of the Free .beta.-Subunit of Choriogonadotropin and Its Potential Value in Screening for Down's Syndrome". J. Endocr. (1980), 84, 295-310, Storring, P. et al. "International Reference Preparation of Human Chorionic Gonadotrophin For Immunoassay: Potency Estimates in Various Bioassay and Protein Binding Assay Systems; And International Reference Preparations of the Alpha and .beta. Subunits for Human Chorionic Gonadotrophin for Immunoassay". Am J. Obstet Gynecol, vol. 155, No. 2, pp. 240-246, Aug. 1986, Macri, J. "Critical Issues in prenatal maternal serum Alpha-fetoprotein screening for genetic anomalies". Am J Obset Gynecol. vol. 148, pp. 241-254, Feb. 1984, Adams et al., "Clinical interpretation of maternal serum alpha-fetoprotein concentrations". Clin. Chem. 36/4, 651-655 (1990), Thomas et al., Human Choriogonadotropin (hCG): Comparisons between Determinations of Intact hCG, Free HCB .beta.-Subunit, "Total" hCG +.beta. in Serum during the First Half of High-Risk Pregnancy. American Journal of Medical Genetics, 36:480-483 (1990), Ozturk et al., "Abnormal Maternal Serum Levels of Human Chorionic Gonadotropin Free Subunits in Trisomy 18". American Journal of Obsetrics and Gynecology, vol. 163, No. 4, pp. 1248-1253, Oct. 1990, Macri et al., "Maternal Serum Down Syndrome Screening: Free .beta.-Protein is a More Effective Marker than Human Chorionic Gonadotropin". Endoctrinology, vol. 126, No. 2, pp. 687-694 (1990), Puisieux et al., "Occurrence of Fragmentation of Free and Combined Forms of the .beta.-Subunit of Human Chorionic Gonadotropin". |
______________________________________ Marker Assay Technique ______________________________________ MSAFP Enzyme linked immunosorbent assay (ELISA) UE Radioimmunoassay Intact HCG Bead type ELISA free beta (HCG) ELISA ______________________________________
______________________________________ FALSE VARIABLE SENSITIVITY POSITIVES ______________________________________ MSAFP 15.4% 4.2% UE 15.4% 2.8% Intact HCG 37% 8.6% MSAFP, UE Intact HCG 50.0% 7.2% free beta (HCG) + Intact HCG 60.0% 8.5% Composite w/o ratio 76% 5.3% Composite w/o UE 76% 5.3% Composite 80% 4.3% Composite w/o free beta HCG 60.0% 5.3% *log intact HCG + 68% 7.6% (log free beta (HCG) + intact HCG) *log intact HCG, log MSAFP 88% 7.4% (log free beta (HCG) + intact HCG) ______________________________________ Composite = MSAFP + free beta (HCG) + Intact HCG + UE + Ratio Gestational age is incorporated along with each variable Risk cutoff level = 1 in 400 except (*) which is 1 in 365.
______________________________________ Marker Assay ______________________________________ MSAFP ELISA (enzyme linked immunosorbent assay) Intact HCG ELISA Intact hcg + ELISA free beta HCG free beta HCG RIA (radio immunoassay) free alpha-HCG RIA UE 2 methods, Enzyme Immunoassay & RIA ______________________________________
______________________________________ False Detection Positive Efficiency Controls Affected ______________________________________ ALL WEEKS Log (beta-1) 6.6% 69.2% 152 26 Log (beta-1) + 5.9% 72.0% 152 25 Log (AFP) Log (beta-2) 8.2% 33.3% 138 18 Log (beta-2) + 10.1% 64.7% 138 17 Log (AFP) Log (beta-2)* 9.6% 33.3% 136 18 Log (beta-2) + 10.3% 52.9% 136 17 Log (AFP)* Weeks 14-16 Log (beta-1) 5.8 68.4% 104 19 Log (beta-1) + 4.8% 73.7% 104 19 Log (AFP) Log(beta-2) 7.1% 45.4% 98 11 Log(beta-2) + 9.2% 63.6% 98 11 Log (AFP) Log (beta-2)* 10.4% 54.6% 96 11 Log (beta-2) + 8.3% 63.6% 96 11 Log (AFP)* ______________________________________ Note: All analyses included gestational age *In the analyses with a * next to them 2 outliers with values of 260 and 316 were removed.